Pseudomonas aeruginosa is a bacterial pathogen which causes infections of the pulmonary tract, urinary tract, burns, wounds, and also causes other blood infections. It is the most common cause of infections of burn injuries and of the external ear (otitis externa), and is the most frequent colonizer of medical devices (e.g., catheters). Pseudomonas can, in rare circumstances, cause community-acquired pneumonias, as well as ventilator-associated pneumonias, being one of the most common agents isolated in several studies. One in ten hospital-acquired infections are from Pseudomonas. 
Detection of P. aeruginosa in clinical settings is still mostly performed by culturing sample bacteria on selective plates or based on immunological tests comprising antibodies to surface antigens of the bacterium, such as outer polysaccharide matrices. Rapid diagnosis and detection is important to start suitable therapy and to control the spread of the bacterium.
Anthrax is a zoonotic disease caused by the spore-forming bacterium Bacillus anthracis. B. anthracis spores remain viable in the environment for years, representing a potential source of infection. Anthrax occurs in humans in three clinical forms: inhalational, gastrointestinal, and cutaneous. Inhalational anthrax results from aerosolization of B. anthracis spores through industrial processing or intentional release. Gastrointestinal or oropharyngeal forms of the disease result from ingestion of infected undercooked or raw meat. Cutaneous anthrax is the most common type of naturally acquired anthrax infection and usually occurs after skin contact with contaminated products from infected animals. Historically, the case-fatality rate for cutaneous anthrax has been <1% with antibiotic treatment and 20% without antibiotic treatment (Brachman P. S. and Kaufmann A. F. In: Evans A. S., Brachman P. S., eds. Bacterial infections of humans. New York: Plenum Medical Book Company, 1998:95-111; Dixon T. C. et al, N. Engl. J. Med. 1999;341:815-26). Case-fatality rates for inhalational anthrax are high, even with appropriate antibiotics and supportive care (Jernigan J. A., et al. Emerg. Infect. Dis. 2001;7:933-44). Among the 18 cases of inhalational anthrax identified in the United States during the 20th century, the overall case-fatality rate was >75%. After the biologic terrorism attack in fall 2001 in which B. anthracis spores were released through the mail, the case-fatality rate for patients with inhalational anthrax was 45% (5 of 11 cases) (Jernigan D. B., et al., Emerg. Infect. Dis. 2002;8:1019-28). The incubation period for anthrax is usually <2 weeks; however, because of spore dormancy and slow clearance from the lungs, the incubation period for inhalational anthrax can be prolonged for months. This phenomenon of delayed onset has not been recognized for cutaneous or gastrointestinal exposures. Discharges from cutaneous lesions are potentially infectious, but person-to-person transmission has been reported rarely. Person-to-person transmission of inhalational anthrax has not been documented.
B. anthracis is one of the biologic agents most likely to be used as a weapon because 1) its spores are highly stable; 2) the spores can infect through the respiratory route; and 3) the resulting inhalational disease has a high case-fatality rate. In 1979 an unintentional release of B. anthracis spores from a military microbiology facility in the former Soviet Union resulted in 69 deaths (Meselson M., et al. Science 1994;266:1202-8). The anthrax outbreak after B. anthracis spores were distributed through the U.S. mail system in 2001 further underscores the dangers of this organism as a terrorist threat.
After a terrorist attack, exposures to B. anthracis spores can occur through primary and secondary aerosols. Primary aerosols are dispersions of particles in air resulting from a biologic agent's initial release, whether through a disseminating device or through handling of an agent-containing package (e.g., in mechanical processing of mail). Secondary aerosols result from disruption and resuspension of settled particles.
Many detection systems for the presence of anthrax have been proposed. Archaic technologies such as staining have nowadays been replaced by more reliable molecular diagnostics, such as PCR (e.g. Makino, S. T. et al., J. Clin. Microbiol. 31:547-51, 1993); U.S. Pat. No. 6,884,588; Qiagen's Real-Art™ B. anthracis PCR) and immunoassays (e.g. Swiecki, M. K., et al. J. Immunol. 176:6076-84, 2006; U.S. Pat. No. 6,828,110; Response Medical Corp.'s RAMP™ Anthrax Assay). However, many of these systems can not differentiate between virulent and avirulent strains of Bacillus anthracis. The virulence of anthrax is mainly given by the presence of the pXO1 plasmid, on which plasmid the gene for the toxic proteins of anthrax are situated, of which one is denominated the lethal factor (LF-protein).
A relatively novel, very robust and highly reliable technology for the visualization of biological material is the FRET (Fluorescent Resonance Energy Transfer) technology. In this process, a photon from an energetically excited fluorophore, the ‘donor’, raises the energy state of an electron in another molecule, the ‘acceptor’, to higher vibrational levels of the excited state. As a result, the energy level of the donor fluorophore returns to the ground state, without emitting fluorescence. The acceptor thus functions as a quencher of the fluorescence. This mechanism is limited by the distance between the donor and the acceptor molecule. Typical effective distances between the donor and acceptor molecules are in the 10 to 100 Å range, which makes them useful in molecular diagnostics of e.g. nucleic acids and proteins.
The FRET technology has been used to detect Bacillus anthracis through coupling of FRET components to nucleic acids, especially those resulting from PCR products (Qi Y. et al., Appl. Environm. Microbiol. 67:3720-7, 2001; Patra G., et al., Annal. New York Acad. Sci. 969:106-11, 2002; Mathur, N. et al., J. Sensors 2008, Art. ID #270475). However, using FRET in combination with PCR still requires a high skill level of the technician performing the assay. Recently, FRET technology has been applied for the detection of anthrax through the proteolytic characteristics of the lethal factor protein (LF). In this assay a labeled substrate was added to the sample which is cleavable by LF (Cummings, R. T. et al., Proc. Natl. Acad. Sci. USA 99:6603-6, 2002). However, this assay gives rise to false positive signals since the cleavage characteristics for the substrate are not specific.
For P. aeruginosa a FRET based system has been described based on DNA (Mancini, n. et al., 2009, J. Clin. Microbiol. doi:10.1128/JCM.00011-09). However, in this assay a DNA lysing step and a PCR step should be performed in order to provide sufficient DNA.
Thus there is still need for an improved detection system for pathogenic microorganisms such as B. anthracis and P. aeruginosa. 